Using a cohort of more than 400 subjects with a wide variety of eosinophilic disorders, ranging from benign eosinophilia to eosinophilic leukemia, we have continued to identify and characterize novel subgroups of patients with eosinophilia. Episodic Angioedema with Eosinophilia (EAE; Gleichs Syndrome) is a rare disorder characterized by episodes of angioedema and eosinophilia that occur at monthly intervals and resolve spontaneously without therapy. Although the etiology of EAE is not yet known, we have demonstrated that multiple lineages, including lymphocytes, neutrophils and mast cells, are involved and may be related to disease pathogenesis (Khoury et al. Haematologica 2015). Whether these cells act directly or promote eosinophilia and eosinophil activation remains to be elucidated. Genetic studies are currently underway. Other subgroups of eosinophilic subjects currently under study include families with autosomal dominant eosinophilia, pediatric patients (Williams et al. J Allergy Clin Immunol 2014:133:AB149, paper submitted), and a cohort of subjects with eosinophilic dermatitis (Khoury et al. J Allergy Clin Immunol 2015:135:AB266). Therapy for eosinophilic disorders remains a primary focus of our group. In the past year, we have reached 50% enrollment in two clinical trials of targeted therapy for the treatment of HES. The first of these is a placebo-controlled, double-blind phase 2 trial of benralizumab (MedImmune/Astra Zeneca), an afucosylated antibody to IL-5 receptor alpha that has shown clinical efficacy in patients with eosinophilic asthma. The current trial stems, in part, from our prior pre-clinical work examining IL-5 receptor levels in patients with eosinophilia and/or mastocytosis (Wilson et al. J Allergy Clin Immunol 2011). To date, 10 of 20 planned subjects have been enrolled, of which 8 have entered the open-label phase of the study. The drug has been well-tolerated and early efficacy results are promising. The second trial is an open-label trial of dexpramipexole (Knopp Biosciences) in patients with steroid-resistant HES. Dexpramipexole was developed for the treatment of amyotrophic lateral sclerosis (ALS). Although it was well tolerated, dexpramipexole failed to show efficacy in a large phase 3 trial for this disease. It did, however, appear to selectively lower blood eosinophils, which prompted the current proof-of-concept open-label phase 2 trial in HES. To date, 5 of the 10 planned subjects have been enrolled. The drug has been well-tolerated and early efficacy results are encouraging. We have also completed enrollment in a new multicenter placebo-controlled, double-blind phase 3 clinical trial of mepolizumab (anti-IL-5 antibody) for the treatment of eosinophilic granulomatosis with polyangiitis (EGPA). In addition to the clinical trials described above, we continue to explore standard therapies for HES. We have analyzed data from an ongoing clinical trial of imatinib in subjects with FIP1L1/PDGFRA-positive myeloproliferative neoplasm (FP), PDGFRA-negative HES with myeloproliferative features (MP) and steroid-resistant without myeloproliferative features (SR) (Khoury et al. presented at the annual meeting of the International Eosinophil Society; manuscript submitted). Overall, imatinib response rates were 100% in the FP group (n= 16), 50% in the MP group (n=13) and 0% in the SR group (n=16). The presence of &#8805; 4 myeloproliferative features was the sole predictor of response. After &#8805; 18 months in complete remission, imatinib was tapered and discontinued in 8 FP and 1 MP subjects. Seven subjects (6 FP, 1 MP) remain in remission off therapy for a median of 29 months (range 14-36). We have also continued to enroll subjects in a double-blind clinical trial designed to assess the utility of a glucocorticoid challenge in determination of glucocorticoid responsiveness in HES and in a clinical trial of mepolizumab (humanized anti-IL-5 antibody) for treatment-refractory HES. Finally, in collaboration with GlaxoSmithKline and ICON, we have begun to collect data for development of a PRO (patient-reported outcome measure) for use in clinical trials in HES. We continue to explore the therapeutic potential of novel antibodies with the potential to target eosinophils, including two antibodies to the inhibitory surface receptors, siglec-8. Siglec-8 is an inhibitory receptor highly expressed on mature eosinophils in blood, bone marrow and tissue. Engagement of the receptor by its ligand or anti-Siglec 8 antibody induces eosinophil apoptosis in the presence of IL-5. Siglec-8 expression was quantified by flow cytometry using whole blood from 38 normal donors (ND) and 58 subjects with HES and was found to be high on eosinophils from normal and HES subjects, irrespective of HES subtype or therapy. Two humanized anti-Siglec-8 antibodies are currently in pre-clinical development, an IgG4 antibody and an IgG1 antibody that has been engineered to cause enhanced ADCC. We have demonstrated that both IgG1 and IgG4 anti-siglec 8 antibodies induce eosinophil apoptosis with similar efficiency in an in vitro assay using whole blood from normal and eosinophilic subjects in the presence of IL-5. In contrast, enhanced ADCC is observed only with the IgG1 antibody and may be limited in patients with very high eosinophil counts and a low NK:EO ratio (Legrand et al. J Allergy Clin Immunol 2015;135:AB221). Murine studies with the two antibodies are currently underway using a humanized IL-2 transgenic NOG mouse model. Helminth infections can be associated with dramatic eosinophilia and complications that are indistinguishable from HES. This is particularly true in loiasis where symptoms are associated with increased eosinophil granule protein levels (indicative of eosinophil activation and degranulation) as well as elevated levels of eosinophil-associated cytokines (Herrick et al. Clin Inf Dis 2015;60:55-63). In some infections, eosinophilia and eosinophil-related symptoms can be exacerbated by effective treatment. For example, treatment of the filarial infection loiasis with diethylcarbamazine (DEC) can be associated with severe adverse events, including death, that are accompanied by a dramatic rise in eosinophilia and are positively correlated with the number of circulating microfilariae in the blood. In the past year, we have enrolled the final subject in a double-blind, placebo-controlled study of the anti-IL-5 antibody, reslizumab, to prevent post-treatment side effects of DEC in patients with Loa loa infection. Finally, in the past year, we have continued to look for biomarkers of eosinophilic disease activity for use in monitoring responses to therapy and helping to dissect out the mechanisms of eosinophil pathogenesis. To this end, we have continued to explore the utility of our multiplex suspension array system developed to simultaneously measure the concentrations of 4 eosinophil granule proteins in serum and plasma (MBP, ECP, EDN and EPO) (Makiya et al. J Immunol Methods 2014) in biological fluids including stool, sputum and urine. Data to date suggest that increased levels of eosinophil granule proteins in stool may be a useful marker of gastrointestinal eosinophilia (Makiya et al. presented at the annual meeting of the International Eosinophil Society 2015) .